The present invention relates to characterization of tissue. More particularly the invention relates to methods of classifying a live subject according to optical thickness of skin, utilizing noninvasive NIR spectroscopy techniques.
Introduction
For many years near infrared (NIR) spectroscopy has been used in the food and agriculture industries to analyze ground wheat and other turbid samples. See P. Williams, K. H. Norris, eds., Near Infrared Technology in the Agricultural and Food Industries, American Association of Food Chemists, St. Paul Minn. (1987). Recently, NIR has found increasing use in biomedical applications, including the nondestructive monitoring of pharmaceuticals and the transcutaneous measurement of analytes in biological tissue. See C. M. Horland, B. Davies, Proc. SPIE 1320, 46 (1990). Also see M. R. Robinson, R. P. Eaton, D. M. Haaland, G. W. Koepp, E. V. Thomas, B. R. Stallard, P. L. Robinson, Clin. Chem., 38:1618-1622 (1992); or J. J. Burmeister, M. A. Arnold, G. W. Small, Diabetes Technology and Therapeutics, 1:5-16 (2000); or S. F. Malin, T. L. Ruchti, T. B. Blank, S. N. Thennadil, S. L. Monfre, Clin. Chem., 45:1651-1658 (1999) 1999; or O. S. Khalil, Clin. Chem., 45:165-177 (1999). All such applications are possible due to the ability of NIR spectroscopy to extract chemical information from complex, highly scattering materials.
Structure of Human Skin
The structure and pigmentation of human skin vary widely among individuals, as well as between different sites on the same individual. Skin consists of a stratified, cellular epidermis, and an underlying dermis of connective tissue. Below the dermis is the subcutaneous fatty layer or adipose tissue. The epidermis is the thin outer layer that provides a barrier to infection and moisture loss, while the dermis is the thick inner layer that provides mechanical strength and elasticity. The epidermis layer is 10-150 xcexcm thick and can be divided into three layers, the basal, middle and superficial layers. The basal layer borders the dermis and contains pigment-forming melanocyte cells, keratinocyte cells, Langherhan cells and Merkel cells. See Ebling, F. J. The Normal Skin. In Textbook of Dermatology, 2nd ed.; Rook, A.; Wilkinson, D. S.; Ebling, F. J. G., Eds.; Blackwell Scientific: Oxford, 1972; pp 4-24. The superficial layer is also known as the stratum corneum.
The stratum corneum, the outermost layer of the mammalian epidermis, is formed and continuously replenished by the slow upward migration of aqueous keratinocyte cells from the germinative basal layer of the epidermis. It is replenished about every two weeks in mature adults. See W. Montagna, The Structure and Function of Skin, 2nd ed., p 454, Academic, New York (1961). This complex process, involving intracellular dehydration and synthesis of an insoluble protein, keratin, results in keratin-filled, biologically inactive, shrunken cells. These flat, dehydrated, hexagonal cells are tightly bound to their neighbors and each is approximately 30 xcexcm wide and 0.8 xcexcm deep. See Baker, H. The Skin as a Barrier. In Textbook of Dermatology, 2nd ed.; Rook, A.; Wilkinson, D. S.; Ebling, F. J. G., Eds.; Blackwell Scientific: Oxford, 1972; pp 249-255. There are about 12 to 20 cell layers over most of the body surface. The stratum corneum is typically 10-20 xcexcm thick, except on the palms and soles, where it is considerably thicker. See A. M. Kligman, The biology of the stratum corneum, in: The Epidermis, W. Montagna, W. C. Lobitz, eds., 387-433 Academic Press, New York (1964).
The major constituent of the dermis, apart from water, is a fibrous protein, collagen, which is embedded in a ground substance composed mainly of protein and glycosaminoglycans. The glycosaminoglycans play a key role in regulating the assembly of collagen fibrils and tissue permeability to water and other molecules. See K. Trier, S. B. Olsen, T. Ammitzboll, Acta. Ophthalmol., 69:304-306 (1999). Collagen is the most abundant protein in the human body. Elastin fibers are also plentiful though they constitute only a small proportion of the bulk. The dermis also contains other cellular constituents, and has a very rich blood supply, though no vessels pass the dermo-epidermal junction. See Ebling, F. J. The Normal Skin. In Textbook of Dermatology, 2nd ed.; Rook, A.; Wilkinson, D. S.; Ebling, F. J. G., Eds.; Blackwell Scientific: Oxford, 1972; pp 4-24. The blood vessels nourish the skin and control body temperature. In humans, the thickness of the dermis ranges from 0.5 mm over the eyelid to 4 mm on the back and averages approximately 1.2 mm over most of the body. See S. B. Wilson, V. A. Spence, Phys. Med. Biol., 33:894-897 (1988).
Optical Properties of Human Skin
When a beam of light beam impinges on the skin, a part of it is reflected, while the remaining part penetrates the skin. The proportion of reflected light energy is strongly dependent on the angle of incidence. At nearly perpendicular incidence, about four percent of the incident beam is reflected due to the change in refractive index between air (xcex7D=1.0) and dry stratum corneum (xcex7D=1.55). For normally incident radiation, this specular reflectance component may be as high as seven percent, because the very rigid and irregular surface of the stratum corneum produces off-normal angles of incidence. Regardless of skin color, specular reflectance of a nearly perpendicular beam from normal skin is always between four and seven percent over the entire spectrum from 250-3000 nm. See J. A. Parrish, R. R. Anderson, F. Urbach, D. Pitts, UV-A: Biologic Effects of Ultraviolet Radiation with Emphasis on Human Responses to Longwave Ultraviolet, Plenum Press, New York (1978). Only the air-stratum corneum border gives rise to a regular reflection. Results from a previous study indicate that the indices of refraction of most soft tissue (skin, liver, heart, etc) lie within the 1.38-1.41 range with the exception of adipose tissue, which has a refractive index of approximately 1.46. See F. P Bolin, L. E. Preuss, R. C. Taylor, R. J. Ference, Appl. Opt., 28:2297-2303 (1989). Therefore, these differences in refractive indices between the different layers of the skin are too small to produce a noticeable reflection. The differences are expected to be even more insignificant when the stratum corneum is hydrated, because of refractive index matching.
The ninety-three to ninety-six percent of the incident beam that enters the skin is attenuated due to absorption or scattering within any of the layers of the skin. These two processes taken together essentially determine the penetration of light into skin, as well as remittance of scattered light from the skin. Diffuse reflectance or remittance is defined as that fraction of incident optical radiation that is returned from a turbid sample. Absorption by the various skin constituents mentioned above accounts for the spectral extinction of the beam within each layer. Scattering is the only process by which the beam may be returned to contribute to the diffuse reflectance of the skin. Scattering results from differences in a medium""s refractive index, corresponding to differences in the physical characteristics of the particles that make up the medium. The spatial distribution and intensity of scattered light depends upon the size and shape of the particles relative to the wavelength, and upon the difference in refractive index between the medium and the constituent particles.
The reduced scattering coefficient of biological tissue depends on many uncontrollable factors, including: concentration of interstitial water, density of structural fibers, and the shapes and sizes of cellular structures. Scattering b y collagen fibers is of major importance in determining the penetration of optical radiation within the dermis. See R. R. Anderson, J. A. Parrish, J. Invest. Dermatol., 77:13-19 (1981). The greater the diffusing power of a medium is, the greater the absorption due to multiple internal reflections. Therefore, reflectance values measured on different sites on the same person, or from the same site on different people, can differ substantially even when the target absorber is present in the same concentration. These differences can be attributed to gender, age, genetics, disease, and exogenous factors due to differences in life styles. For example, it is known that, in humans, skin thickness is greater in males than in females, whereas the subcutaneous fat thickness is greater in females. The same group reports that collagen density, the packing of fibrils in the dermis, is higher in the forearms of males than females. See S. Schuster, M. M. Black, E. McVitie, Br. J. Dermatol., 93:63-643 (1975). Due to the wide variation in the amount of collagen present in the dermal layer per unit of volume, physical thickness of the skin is not strictly correlated to the amount of collagen in the dermal layer. Thus, geometric skin thickness alone is not a reliable predictor of the optical properties of an individual subject""s skin.
The scattered light remitted from the skin is a mixture of short-path backscattered photons and long-path diffusively scattered photons. The proportion of each is dependent on the separation between the source and detector probes of the spectroscopic instrument. Short-path, backscattered photons dominate the spectral information at small source-detector separations, while long-path, diffusely scattered photons dominate at large source-detector separations. At points closer to the source, the backscattered photons have spent a smaller fraction of their total optical path propagating through the turbid media. At points further from the source, more photons undergo multiple scattering events, and the fraction of their total optical paths spent in the deeper turbid tissue becomes greater. Hence, spectra at large source-detector separations are expected to be heavily weighted by features that correspond to absorbing species located deeper in the tissue. See G. Kumar, J. M Schmitt, Appl. Opt. 36:2286-2293 (1997). In summary, for heterogeneous tissue, the spectral characteristics of diffuse remittance are the result of interaction of;
(1) absorption and scattering properties of the tissue;
(2) the distribution of absorbing species and scattering components; and
(3) the source-tissue-detector geometry.
Thus, it would be desirable to adapt probe geometry to maximize the collection of light that has been diffusely reflected from a desired depth or depth range in the skin. Furthermore, it would be a great advantage to classify subjects according to the optical properties of their skin, thus facilitating the screening of those subjects whose skin composition does not match the probe specifications.
The present invention provides a method for assessment and classification of a subject according to the optical thickness of the skin as determined by noninvasive, near-infrared reflectance measurements on skin tissue. The assignment of this optical skin thickness characteristic is performed by an analysis of the water, fat, and protein marker bands in the near infrared spectrum. The optical skin thickness characteristic is then used to evaluate the suitability of the subject for calibration on a standardized set of optical probes. The optical probes in the standardized set are designed to cover a range of penetration depths by varying the distance distribution between illuminator and detector fibers on each probe tip.
The invention is not limited to the classification of subjects based on the optical thickness of their skin for the noninvasive measurement of blood analytes, but also finds application in diagnosis and investigation of diseases in which a generalized defect of connective tissue is suspected. The measurement of skin thickness, collagen content and density also provides useful information in metabolic and endocrine diseases. See C. Hamlin, R. R. Kohn, J. H. Luschin, Diabetes, 24:902-904 (1975) or S. L. Schnider, R. R. Kohn, J. Clin. Invest., 66:1179-1181 (1980).